• Title/Summary/Keyword: defense-related enzyme

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Molecular Characterization of a thiJ-like Gene in Chinese Cabbage

  • Oh, Kyung-Jin;Park, Yong-Soon;Lee, Kyung-Ah;Chung, Yong-Je;Cho, Tae-Ju
    • BMB Reports
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    • v.37 no.3
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    • pp.343-350
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    • 2004
  • A cDNA clone for a salicylic acid-induced gene in Chinese cabbage (Brassica rapa subsp. pekinensis) was isolated and characterized. The cabbage gene encoding a protein of 392 amino acids contained a tandem array of two thiJ-like sequences. ThiJ is a thiamin biosynthesis enzyme that catalyzes the phosphorylation of hydroxymethylpyrimidine (HMP) to HMP monophosphate. Although the cabbage gene shows a similarity to bacterial thiJ genes, it also shares a similarity with the human DJ-1, a multifunctional protein that is involved in transcription regulation, male fertility, and parkinsonism. The cabbage thiJ-like gene is strongly induced by salicylic acid and a nonhost pathogen, Pseudomonas syringae pv. tomato, which elicits a hypersensitive response in Chinese cabbage. Treatment of the cabbage leaves with BTH, methyl jasmonate, or ethephon showed that the cabbage thiJ-like gene expression is also strongly induced by BTH, but not by methyl jasmonate or ethylene. This indicates that the cabbage gene is activated via a salicylic acid-dependent signaling pathway. Examination of the tissue-specific expression revealed that the induction of the cabbage gene expression by BTH occurs in the leaf, stem, and floral tissues but not in the root.

Studies on the Regulation of Nitric oxide Synthesis in Murine Mononuclear Phagocytes (마우스 단핵 탐식 세포에서 Nitric oxide 생성의 조절 기전에 관한 연구)

  • 최병기;김수응
    • Environmental Analysis Health and Toxicology
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    • v.15 no.3
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    • pp.69-80
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    • 2000
  • ADP-rubosylation may be involved in the process of macrophage activation. Nitric oxide (NO) has emerged as an important intracellular and interacellular regulatory molecule with function as diverse as vasodilation, neural communication or host defense. NO is derived from the oxidation of the terminal guanidino nitrogen atom of L-arginine by the NADPH -dependent enzyme, nitric oxide synthase (NOS) which is one of the three different isomers in mammalian tissues. Since NO can exert protective or regulatory functions in the cell at a low concentration while toxic effects at higher concentrations, its role may be tightly regulated in the cell. Therefore, this paper was focused on signal transduction pathway of NO synthesis, role of endogenous TGF-$\beta$ in NO production. effect of NO on superoxide formation. Costimulation of murine peritoneal macrophages with interferon-gamma (IFN-γ) and phorbol 12-myristate 13-acetate (PMA) increased both NO secretion and mRNA expression of inducible nitric oxide synthase (iNOS) when PMA abolished costimulation. Pretreatmnet of the cells with PMA abolished costimuation effects due to the depletion of protein kinase C (PKC) activities . The involvement of PKC in NO secretion could be further confirmed by PKC inhibitor, stauroprine, and phorbol ester derivative, phorbol 12,13-didecanoate. Addition of actinomycine D in IFN-γ plus PMA stimulated cells inhibited both NO secretion and mRNA expression of iNOS indication that PMA stabilizes mRNA of iNOS . Exogenous TGF-$\beta$ reduced NO secretion in IFN -γ stimulated murine macrophages. However addition of antisense oligodeoxynucleotide (ODN) to TGF-$\beta$ to this system recovered the ability of NO production and inhibited mRNA expression of TGF-$\beta$. ACAS interactive laser cytometry analysis showed that transportation of FITC -labeled antisense ODN complementary to TGF-$\beta$ mRNA could be observed within 5 min and reached maximal intensity in 30 min in the murine macrophage cells. NO released by activated macrophages inhibits superoxide formation in the same cells . This inhibition nay be related on NO-induced auto -adenosine diphosphate (ADP) -ribosylation . In addition, ADP-ribosylation may be involved in the process of macrophage activation .

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Identification of another calmodulin-binding domain at the C-terminal region of AtCBP63

  • Kim, Sun-Ho;Kang, Yun-Hwan;Han, Hay-Ju;Bae, Dong-Won;Kim, Min-Chul;Lim, Chae-Oh;Chung, Woo-Sik
    • Journal of Plant Biotechnology
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    • v.36 no.1
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    • pp.53-58
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    • 2009
  • Calcium signals can be transduced by binding calmodulin (CaM), a $Ca^{2+}$ sensor in eukaryotes, is known to be involved in the regulation of diverse cellular functions. We isolated a CaM-binding protein 63 kD (AtCBP63) from the pathogen-treated Arabidopsis cDNA expression library. Recently, AtCBP63 was identified as a CaM bining protein. The CaM binding domain of AtCBP63 was reported to be located in its N-terminal region, In this study, however, we showed that ACaM2 could specifically bind to second CaM-binding domain (CaMBD) of AtCBP63 at the C-terminal region. The specific binding of CaM to CaM binding domain was confirmed by a gel mobility shift assay, a split ubiquitin assay, site-directed mutagenesis, and a competition assay using a $Ca^{2+}$/CaM-dependent enzyme. The gene expression of AtCBP63 was induced by pathogens and pathogens related second messengers. This result suggests that a CaM binding protein, AtCBP63, may play role in pathogen defense signaling pathway.

Protein-protein Interaction Network Analyses for Elucidating the Roles of LOXL2-delta72 in Esophageal Squamous Cell Carcinoma

  • Wu, Bing-Li;Zou, Hai-Ying;Lv, Guo-Qing;Du, Ze-Peng;Wu, Jian-Yi;Zhang, Pi-Xian;Xu, Li-Yan;Li, En-Min
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.5
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    • pp.2345-2351
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    • 2014
  • Lysyl oxidase-like 2 (LOXL2), a member of the lysyl oxidase (LOX) family, is a copper-dependent enzyme that catalyzes oxidative deamination of lysine residues on protein substrates. LOXL2 was found to be overexpressed in esophageal squamous cell carcinoma (ESCC) in our previous research. We later identified a LOXL2 splicing variant LOXL2-delta72 and we overexpressed LOXL2-delta72 and its wild type counterpart in ESCC cells following microarray analyses. First, the differentially expressed genes (DEGs) of LOXL2 and LOXL2-delta72 compared to empty plasmid were applied to generate protein-protein interaction (PPI) sub-networks. Comparison of these two sub-networks showed hundreds of different proteins. To reveal the potential specific roles of LOXL2- delta72 compared to its wild type, the DEGs of LOXL2-delta72 vs LOXL2 were also applied to construct a PPI sub-network which was annotated by Gene Ontology. The functional annotation map indicated the third PPI sub-network involved hundreds of GO terms, such as "cell cycle arrest", "G1/S transition of mitotic cell cycle", "interphase", "cell-matrix adhesion" and "cell-substrate adhesion", as well as significant "immunity" related terms, such as "innate immune response", "regulation of defense response" and "Toll signaling pathway". These results provide important clues for experimental identification of the specific biological roles and molecular mechanisms of LOXL2-delta72. This study also provided a work flow to test the different roles of a splicing variant with high-throughput data.

Induction of Systemic Resistance against Cucumber mosaic virus in Arabidopsis thaliana by Trichoderma asperellum SKT-1

  • Elsharkawy, Mohsen Mohamed;Shimizu, Masafumi;Takahashi, Hideki;Ozaki, Kouichi;Hyakumachi, Mitsuro
    • The Plant Pathology Journal
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    • v.29 no.2
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    • pp.193-200
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    • 2013
  • Trichoderma asperellum SKT-1 is a microbial pesticide that is very effective against various diseases. Our study was undertaken to evaluate T. asperellum SKT-1 for induction of resistance against yellow strain of Cucumber mosaic virus (CMV-Y) in Arabidopsis plants. Disease severity was rated at 2 weeks post inoculation (WPI). CMV titre in Arabidopsis leaves was determined by indirect enzyme-linked immunosorbent assay (ELISA) at 2 WPI. Our results demonstrated that among all Arabidopsis plants treated with barley grain inoculum (BGI) of SKT-1 NahG and npr1 plants showed no significant reduction in disease severity and CMV titre as compared with control plants. In contrast, disease severity and CMV titre were significantly reduced in all Arabidopsis plants treated with culture filtrate (CF) of SKT-1 as compared with control plants. RT-PCR results showed increased expression levels of SA-inducible genes, but not JA/ET-inducible genes, in leaves of BGI treated plants. Moreover, expression levels of SA- and JA/ET-inducible genes were increased in leaves of CF treated plants. In conclusion, BGI treatment induced systemic resistance against CMV through SA signaling cascade in Arabidopsis plants. While, treatment with CF of SKT-1 mediated the expression of a majority of the various pathogen related genes, which led to the increased defense mechanism against CMV infection.

Effect of Simulated Acid Rain on Antioxidants and Related Enzymes in Garden Balsam (Impatiens balsamina L.) (봉선화(Impatiens balsamina L.)에 대한 pH 수준별 처리가 항산화 물질 및 관련 효소 활성에 미치는 영향)

  • Kim Hak Yoon
    • Journal of Life Science
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    • v.15 no.3 s.70
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    • pp.382-386
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    • 2005
  • To investigate the effects of simulated acid rain (SAR) on growth and biochemical defense responses of plant, garden balsam (Impatiens balsamina L.) was subjected to four levels of SAR based on pH (5.6, 4.0, 3.0, 2.0) and placed in the growth chambers for 2 weeks. SAR drastically inhibited both height and dry weight of garden balsam. The content of total carotenoid was tended to decrease, but the level of malondialdehyde was significantly increased by SAR. As the pH levels decreased from 5.6 to 2.0, the content of dehydroascorbate and oxidized glutathione of the plant were significantly increased. The enzyme (superoxide dismutase, ascorbate peroxidase etc.) activities of the plant affected by SAR were increased as the pH decreased. The results indicate that garden balsam may receive oxidative stresses by the application of SAR and by which the plant growth can be significantly retarded. A biochemical protective mechanism might be activated to nullify the oxidative stresses generated through SAR.

Immune Enhancing Effects of Pyropia yezoensis Hydrothermal Extract in RAW 264.7 Cells (방사무늬 김 열수추출물의 RAW 264.7 세포에서의 면역 증진 효과)

  • Goeun Jang;Bo-Ram Park;Seul Ah Lee;Chun Sung Kim
    • Journal of Marine Bioscience and Biotechnology
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    • v.15 no.2
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    • pp.33-40
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    • 2023
  • This study aimed to investigate the immunomodulatory function of Pyropia yezoensis hydrothermal (water) extract (PYWE) in comparison to the group treated only with lipopolysaccharides (LPS) in RAW264.7 cells. LPS is known to be an inflammatory mediator that activates macrophages, leading to the secretion of nitric oxide (NO), inducible nitric oxide synthase (iNOS), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) as defense responses. Through enzyme-linked immunoassay and western blot analyses, it was observed that PYWE increased the expression levels of NO, iNOS, TNF-α, and IL-6 in RAW264.7 cells in a dose-dependent manner, although to a lesser extent compared with the group treated with LPS alone. In addition, the study examined the mitogen-activated protein kinases (MAPKs) pathway, which regulates various cellular activities, including gene expression, mitosis, cell differentiation, transformation, survival, and death. The western blot analysis confirmed that PYWE also regulated the MAPKs pathway. Furthermore, the expression levels of immunomodulatory-related factors increased in the group treated with PYWE compared with the control group. Even though the effects of PYWE were usually less strong than those of LPS, the effects of PYWE increased with increasing doses compared to the control group. This suggests that PYWE could be used to control the immune system.

Characterization of SID2 that is required for the production of salicylic acid by using β-GLUCURONIDASE and LUCIFERASE reporter system in Arabidoposis (리포트 시스템을 이용한 살리실산 생합성 유전자 SID2의 발현 해석)

  • Hong, Mi-Ju;Cheong, Mi-Sun;Lee, Ji-Young;Kim, Hun;Jeong, Jae-Cheol;Shen, Mingzhe;Ali, Zahir;Park, Bo-Kyung;Choi, Won-Kyun;Yun, Dae-Jin
    • Journal of Plant Biotechnology
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    • v.35 no.3
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    • pp.169-176
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    • 2008
  • Salicylic acid(SA) is a phytohormone that is related to plant defense mechanism. The SA accumulation is triggered by abiotic and biotic stresses. SA acts as a signal molecular compound mediating systemic acquired resistance and hypersensitive response in plant. Although the role of SA has been studied extensively, an understanding of the SA regulatory mechanism is still lacking in plants. In order to comprehend SA regulatory mechanism, we have been transformed with a SID2 promoter:GUS::LUC fusion construct into siz1-2 mutant and wild plant(Col-0). SIZ1 encodes SUMO E3 ligase and negatively regulates SA accumulation in plants. SID2(SALICYLIC ACID INDUCTION DEFICIENT2) is a crucial enzyme of SA biosynthesis. The Arabidopsis SID2 gene encodes isochorismate synthase(ICS) that controls SA level by conversion of chorismate to isochorismate. We compared the regulation of SID2 in wild-type and siz1-2 transgenic plants that express SID2 promoter:GUS::LUC constructs respectively. The expressions of $\beta$-GLUCURONIDASE and LUCIFERASE were higher in siz 1-2 transgenic plant without any stress treatment. SID2 promoter:GUS::LUC/siz1-2 transgenic plant will be used as a starting material for isolation of siz1-2 suppressor mutants and genes involved in SA-mediated stress signaling pathway.

Effects of Different UV-B Levels on Growth, Antioxidant Contents and Activities of Related Enzymes in Cucumber(Cucumis sativus L.). (UV-B 강도 변화가 오이의 생장 및 항산화 물질 함량과 관련 효소의 활성에 미치는 영향)

  • Kim, Hak-Yoon;Shin, Dong-Hyun;Kim, Kil-Ung
    • Korean Journal of Environmental Agriculture
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    • v.19 no.4
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    • pp.309-313
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    • 2000
  • To investigate the effects of different UV-B levels on growth and biochemical defense response in plants, cucumber plants were subjected to three levels of biologically effective ultraviolet-B $(UV-B_{BE})$ radiation [daily dose: 0.03 (No), 6.40 (Low) and $11.30\;(High)\;kJ{\cdot}m^{-2}$, $UV-B_{BE}$] in the growth chambers for 3 weeks during the early growth period. Enhanced UV-B radiation drastically decreased both dry weight and leaf area of cucumber. With increasing UV-B intensity, chlorophyll content was decreased, however the level of malondialdehyde was highly increased linearly. Total contents of ascorbic acid and glutathione were tended to increase by UV-B, while the ratios of dehydroascorbate/ascorbate and oxidized glutathione/reduced glutathione were significantly increased with increasing UV-B intensity in cucumber. All the enzyme activities investigated (superoxide dismutase, ascorbate peroxidase, dehydroascorbate reductase, guaiacol peroxidase etc.) in cucumber were increased by the UV-B enhancement. These results suggested that enhanced UV-B irradiation caused photooxidative stress in cucumber plant and resulted in significant reduction in plant growth. Biochemical protection responses might be activated to prevent the leaves from damaging effects of oxidative stress generated by UV-B irradiation.

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The Induction of Secondary Toxic Substances and Antioxidative Enzymes by $SO_2$ Fumigation in Foliage Plants (관엽식물에서 $SO_2$ 처리에 의한 이차독성물질 생성 및 항산화효소의 유도)

  • Lee, Young-Yi;Kang, Youn-Ji;Lee, Yong-Beom
    • Korean Journal of Environmental Agriculture
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    • v.20 no.1
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    • pp.44-49
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    • 2001
  • To study whether the secondary toxic substances such as ethylene and reactive oxygen species(ROS) are induced by air pollutants in foliage plants, $SO_2$ was fumigated to Pachira aquatica, Spathiphyllum patinii, and Hedera helix. $SO_2$ was controlled to $1\;{\mu}L/L$ and then fiumigated to plants for 2 days(8 hrs/day). It resulted in visible injury in P. aquatica and H. helix while no symptom appeared in S. patinii. Photosynthetic rate and water use efficiency were most remarkably reduced in P. aquatica compared to other two species whereas least in S. patinii. Two days after $SO_2$ fumigation, ethylene evolution was quantified to 23.56, 10.43 and 4.79 nL/g/h in P. aquatica, H. helix and S patinii, respectively. On the other hand, antioxidative enzymes were clearly activated by $SO_2$ treatment in all tested plant species implying ROS production. In conclusion, we could suggest that ethylene and ROS have been intimately related to the defense mechanism against $SO_2$ and their induction degree increased with plant susceptibility to $SO_2$. Furthermore, it was found that S. patinii was tolerant and P. aquatica sensitive to $SO_2$ on the basis of antioxidative enzyme activity and ethylene evolution.

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